Application of two-dimensional reversed phase countercurrent chromatography × high-performance liquid chromatography to bioactivity-guided screening and isolation of α-glucosidase inhibitors from Rheum palmatum L.

In the present work, comprehensive two-dimensional reversed-phase countercurrent chromatography × reversed-phase liquid chromatography combined (2D RPCCC × RPLC) with 2D microfraction bioactive evaluation was employed to screen and isolate α-glucosidase inhibitors from Rheum palmatum L. Countercurrent chromatography was employed to improve 2D analysis and preparative separation. A selected biphasic solvent system composed of petroleum ether/ethyl acetate/methanol/water with gradient elution mode was used for the first dimension RPCCC separation (1D RPCCC). Solid-phase extraction was applied to eliminate interfering polar compounds before the second dimension analysis (2D RPLC). 76 components were shown in 2D contour plot in UV 280 nm. 11 Candidates were separated by a scaled-up CCC and identified by 1H NMR and 13C NMR, including anthraquinones, flavonoids, stilbenes, phenols, and glucoside derivatives. In addition, it was found that two components, resveratrol-4'-O-(6″-galloyl)glucoside (36) and lyciumaside (43) were identified as natural α-glucosidase inhibitors in Rheum palmatum L. for the first time.

Screening and identification of antibacterial components in Artemisia argyi essential oil by TLC-direct bioautography combined with comprehensive 2D GC × GC-TOFMS.

One of the primary components that contribute to Artemisia argyi 's effectiveness is essential oil, which has an exceptional antibacterial effect that has been well documented. The actual cause of its antibacterial activity and associated mechanism, however, are still not completely understood. For the first time, comprehensive two-dimensional gas chromatography coupled with time-of-flight mass spectrometry (2D GC × GC-TOFMS) and thin-layer chromatography-direct bioautography (TLC-DB) were employed to investigate its antibacterial components. The antibacterial properties of A. argyi essential oil were investigated, and the antibacterial activity of six compounds was evaluated, using Staphylococcus aureus (S. aureus) and Escherichia coli (E. coil) as test microorganisms. TLC-direct bioautography was used to screen two bioactive clusters. Following 2D GC × GC-TOFMS identification of bioactive clusters, six compounds were evaluated for in vitro antibacterial activity verification. All the components tested displayed antibacterial action. Results showed that α-terpineol and eugenol had high potent antibacterial activity (MIC＜0.62 mg/mL, IC50＜2.00 mg/mL). For complex essential oils from traditional Chinese medicine, this method is efficient for quick screening and identifying antibacterial compounds.

High-resolution monophenolase/diphenolase/radical scavenging profiling for rapid screening of natural whitening candidates from Rosa rugosa cv. 'Plena'.

Antioxidants and tyrosinase inhibitory components were successfully screened and separated from Rosa rugosa cv. 'Plena' by high-performance liquid chromatography microfractionation bioactive screening combined with several separation and purification methods. Ethyl acetate extract of Rosa rugosa cv. 'Plena' showed high antioxidant activity and tyrosinase inhibitory activity. High-speed countercurrent chromatography, silica gel column chromatography, and semi-preparative high-performance liquid chromatography were used for the preparative separation of four bioactive components from ethyl acetate extract. Two tyrosinase-inhibiting active substances, flavogallonic acid, and N1 -N5 -N10 -tri-4-p-coumaroylspermidine, were isolated from Rosa rugosa cv. 'Plena', and they showed great monophenolase inhibition activity (half-maximal inhibitory concentration: 664.60 and 23.77 µg/ml, respectively) and excellent diphenolase inhibition activity (half-maximal inhibitory concentration: 23 614.61 and 16.80 µg/ml, respectively). Meanwhile, gallic acid, flavogallonic acid, and ellagic acid were shown to have excellent 1,1-diphenyl-2-picryl-hydrazyl antioxidant activity (half maximal inhibitory concentration: 6.66, 20.17, and 13.45 µg/ml), and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) antioxidant activity (half maximal inhibitory concentration: 3.53, 3.83, and 2.78 µg/ml). Molecular docking revealed that flavogallonic acid and N1 -N5 -N10 -tri-4-p-coumaroylspermidine had a strong binding affinity (-9.3 and -10 kcal/mol, respectively) to tyrosinase through hydrogen bonding and hydrophobic interactions.

Comprehensive two-dimensional countercurrent chromatography × gas chromatography characterization of Artemisia argyi essential oil.

A comprehensive two-dimensional (2D) countercurrent chromatography (CCC) × gas chromatography (GC) was investigated for characterization of chemical constituents of Artemisia argyi essential oil, and orthogonality for the 2D chromatographic system was evaluated. A solvent system composed of n-hexane/acetonitrile/methanol (2:2:1, v/v/v) was selected for first dimensional separation of Artemisia argyi essential oil. Then all CCC fractions were analyzed by GC, which provided a wealth of information regarding the composition of the essential oil. Visualization of chemical compositions obtained from the comprehensive 2D CCC × GC separation was achieved by creation of a 2D contour plot map. Total peak capacity was evaluated and approximately 1392 peaks were obtained through a comprehensive 2D CCC × GC separation. A high spatial coverage and a low linear correlation coefficient were achieved. Meanwhile, all compounds were identified by GC-MS. The obtained 2D contour plot could be divided into six zones to show the characteristic chemical compositions. Six zones could be divided into different component groups, including monoterpenes, sesquiterpenes, monoterpene alcohols, phenols, aldehydes, ketones and esters, which could be used to identify compounds that have not been reported, and to predict the structure of unknown compounds in Artemisia argyi essential oil and comprehensively characterize fingerprint peak.

Two-dimensional chromatography in screening of bioactive components from natural products.

INTRODUCTION: Screening and analysis of bioactive components from natural products is a fundamental part of new drug development and innovation. Two-dimensional (2D) chromatography has been demonstrated to be an effective method for screening and preparation of specific bioactive components from complex natural products. OBJECTIVE: To collect details of application of 2D chromatography in screening of natural product bioactive components and to outline the research progress of different separation mechanisms and strategies. METHODOLOGY: Three screening strategies based on 2D chromatography are reviewed, including traditional separation-based screening, bioactivity-guided screening and affinity chromatography-based screening. Meanwhile, in order to cover these aspects, selections of different separation mechanisms and modes are also presented. RESULTS: Compared with traditional one-dimensional (1D) chromatography, 2D chromatography has unique advantages in terms of peak capacity and resolution, and it is more effective for screening and identifying bioactive components of complex natural products. CONCLUSION: Screening of natural bioactive components using 2D chromatography helps separation and analysis of complex samples with greater targeting and relevance, which is very important for development of innovative drug leads.